When cells divide, they must first copy their genome to ensure new cells have all the correct genes and information they need to function properly. The maize genome is almost as large as the human genome, with all that information crammed into just 10 chromosomes (compared to 23 in humans). If all this DNA was replicated at the same time, it wouldn’t be able to fit within the nucleus of the cell. How maize achieves this feat of efficiency and function is central to new research findings recently published in the journal Plant Cell. 

The project began over 20 years ago at NC State University with Randall Shultz, a graduate student in the lab of Bill Thompson, a professor in the Department of Plant and Microbial Biology. Shultz took a biochemistry course taught by Linda Hanley-Bowdoin in which students wrote their own research proposals. He proposed adapting techniques used to explore when sections of the genome are replicated in yeast, and applying them to plants. 

“I wrote on top of the proposal … somebody ought to do this, go talk to Bill. And that was the beginning,” says Hanley-Bowdoin, professor in the Department of Plant and Microbial Biology and principal investigator of the project’s most recent National Science Foundation grant. 

Digging into Plant DNA

The researchers started by characterizing when different parts of the genome were replicated. They found that plants replicate their genomes in smaller segments than animals. This most recent publication takes their work a step further, looking at not only when but also where DNA  replication is occurring in the nucleus. 

“This is the first time anyone has analyzed plant DNA replication at this level of structural detail,” Thompson explains. 

 For the latest project, Hanley-Bowdoin and Thompson partnered with researchers, including several former trainees, Hank Bass at Florida State University, and Lorenzo Concia at the Texas Advanced Computing Center. They focused on corn (Zea mays) root tips. The root tip is an important region because cells there divide rapidly as the root grows deeper into the soil. And it’s a good place to study DNA replication because all those dividing cells need to replicate their DNA.

In the lab at NC State, supervisor Emily Wear and researcher Leigh Mickelson-Young collected isolated nuclei from rapidly growing cells at the tips of maize roots. They first separated the nuclei according to their stage of DNA replication.The sorted nuclei were used to examine where DNA was being replicated at various times during the replication process. 

The researchers labeled the replicating DNA in these samples with a fluorescent “tag” that can be seen through a special microscope. The tag, together with additional molecular analyses, tracked the fate of individual DNA sequences, creating a picture of the replicating DNA within the nucleus. Analyzing such pictures, collaborators Hank Bass and Sara Akram from Florida State University, together with Lorenzo Concia from the Texas Advanced Computing Center, created a three-dimensional model of DNA replication in the maize nucleus.

“They were able to show that the replication patterns look different than they do in animal cells,” Thompson explains. “It’s a story of things being more mixed up, more closely interspersed —  but they could tell the difference between early and middle [replication].” 

Replicating Success

The patterns they saw matched up with earlier work that looked at how densely packed the maize DNA was in the chromosomes. When DNA is densely packed, it’s difficult to access and is less frequently transcribed. More open DNA is associated with more transcription and gene activity and is replicated earlier 

The work in this paper has exciting implications for biotechnology. When sections of the genome replicate, they are more open and accessible, facilitating easier gene editing or  the addition of large cassettes of genes. Future work on molecular and genetic controls of replication timing may help plant breeders manipulate the genome to enhance or create new traits. 

As Hanley-Bowdoin puts it, “The opportunities are starting to become more apparent. And I think the link between replication timing and the three-dimensional structure of chromatin is going to be very valuable information down the road.”

This post was originally published in College of Agriculture and Life Sciences News.